azo dye biobegredation

7/29/2019 azo dye biobegredation

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Presented by

Sabaina TahirM.Phil 3rd

Determination and Biodegradation of

azo dyes in soils contaminated with

textile dye effluent


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Azo Dyes

Azo compounds, i.e. molecules with one or more azo

(N=N) bridges linking substituted aromatic structures

They are generally aromatic hydrocarbons, derivatives of

benzene, toluene, naphthalene, phenol and aniline

Azo dyes are the most important group of syntheticcolorants that are extensively used in textile, food,

pharmaceutical and printing industries


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Physicochemical processes for biodegradation have

limitations and also produce toxic by products Biodegradation using microorganisms is gaining

importance as it is cost effective, environmentally

friendly, and produces less sludge

Different taxonomic groups of bacteria have beenreported for their ability to decolorize azo dyes

Aerobic degradation of azo dyes has been reported by

many investigators

This work has also been undertaken in order toinvestigate the potential of aerobic bacteria to degrade

azo dyes


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Sampling

Soils affected by azo-dye contamination around textile

industries are to be selected for sampling

Soil samples are to be collected from the areas of

Rawalpindi and Gujranwala

Scalpel will be used for this purpose by removing soil layerupto 15cm

Samples are to be stored in polythene bags aerobically at

4C


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Enrichment culture techniques

Enrichment techniques is carried out by taking clay loam

soil in a petri dish and calculated quantity of textile

effluent like azo dyes is added so that, the finalconcentration is around 500 ppm of soil.

Enough quantity of water is added to the petri dish so

that soil becomes moist, stirred well with glass rod andthe Petri plate is covered with lid and incubated for period

of 25 days.


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A solution will be prepared by taking five

grams of enriched soil samples and

transferred to a 250 mL conical flask

containing 100 mL of sterile distilled water,stirred well and the supernatant was diluted

up to five dilutions through a serial

dilution method.


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Isolation

For the dilution plate method 10 g of sample is transferred in 90

mL of 0.85% saline water

Pasteurization at 80C for 10 min

1 mL aliquot from each of the samples is transferred in 9 mL of

0.85% saline water and preparation of 6 fold dilutions as done

1 mL of dilutions was plated on nutrient agar plates/ mineral salt

medium and incubated for 48 - 72 h at 50C.

The plates were covered with aluminium foil and single colonieswith different morphologies were picked and purified using streak

plate method


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Screening

The bacterial isolates are to be sequentially

adapted to higher concentration of azo

dyes by repeated sub culturing on solid

culture media (Mineral salt medium andagar)

The maximum resistance limits (MRL)

beyond which the bacterial growth isdetermined apparent, decolorization

abilities of the bacterial strains are

monitored at 37C for 8 days


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Preservation of isolates

Glycerol stocks are prepared and stored at -

80C for long term preservation

Pure cultures strains are incubated at 50C

for 48 h in isolation broth

Then 0.5 mL of each of the cultures aretransferred into cryotubes with 0.5 mL broth

containing 40% glycerol


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Degradation studies

The isolates obtained are to be placed on Mineral Salt

Medium with different conditions

Growth of isolates is to be observed by incubation at

different temperatures, providing varying pH and with

different substrate concentrations

In all above cases growth of bacterial strains is to be

determined with an interval of 4 days


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Development of consortium in 10 ml culture tubes containingmixture of reactive azo dyes, 5 ml nutrient broth and a loop ful

of each microbial culture (incubated for 24 hrs)

5 ml of each bacterial strain culture is to be added to 250 mL

Erlenmeyer flask containing 100 mL of textile effluent The flask was further incubated to observe the time required for

decolorization

3ml of aliquots of the culture media are to be withdrawn at

different time intervals, centrifuged at 15 min to separate the

bacterial cell mass

Decolorization of the textile effluent was analyzed using a

UV/Vis spectrophotometer (deminishing of peaks in visible

region)


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Morphological characters of the isolates were

determined by gram staining, spore staining and

motility testing

16S rRNA sequencing is to be performed to find out

the nucleotide sequence and it is helpful for

determining the genus of a particular specie


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Thank You

azo dye biobegredation

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